Senescence was initially defined as the stable cell cycle arrest that accompanies replicative exhaustion of human fibroblasts in culture, and may be important in tumor suppression and aging. In addition to telomere malfunction, which induces replicative senescence, a phenotypically identical endpoint can be induced acutely in young cells by a variety of stresses. For example, we have shown that 'premature' cellular senescence (or STASIS) can be induced acutely in various cell types by misexpression of oncogenic ras or DNA damaging agents such as chemotherapeutic drugs. Given these observations, we proposed that senescence parallels apoptosis as a cellular response to stress, and plays similar roles in suppressing tumorigenesis and modulating chemotherapy. Over the course of the previous funding period, we developed a better understanding of the senescence program and how the process is controlled as part of a complex tumor suppressor network. Our work has evolved from identifying upstream signaling pathways that trigger senescence (e.g. MAPK signaling and ARF) to characterizing genes and processes that act further downstream to produce the senescence endpoint (e.g. PML, Rb, and SAHFs). In our renewal application, the effector mechanisms of senescence (i.e. the senescence machinery) will receive the most attention, in part, because very little is known about these processes and because they are controlled by genes with established roles in tumor suppression. Specifically, we will further dissect the role of p53 and Rb in cellular senescence, explore the effector mechanisms of senescence that contribute to its stability, and identify genes that can reverse or bypass the program. Our approach will continue to apply genetic principles to the analysis of cellular senescence in primary fibroblasts and epithelial cells, but will also incorporate expression array, genome wide RNA interference, and proteomics technology to understand the senescence machinery. These studies will provide new insights into the nature of the senescence program and, as such, will further assemble an important tumor suppressor network that provides an important brake to cancer development and may also contribute to age-related diseases. Our studies also may identify new 'biomarkers' of senescence that can eventually be used to study the process in vivo. [unreadable] [unreadable] [unreadable]